Hybrid synapse formation between spinal motoneurons and superior cervical ganglion neurons in vitro: A study of the functional reconstruction of visceral organs

Spinal motoneurons (SMNs) were co-cultured with labeled superior cervical ganglion neurons (SCGNs) in complete medium on coverslips prepared by the microisland technique. “Hybrid” synapses formation between SMNs and SCGNs was examined by double immunostaining, reverse transcription-polymerase chain reaction (RT-PCR), and electrophysiological assays. The number of synaptic puncta was significantly higher in co-cultured cells compared to controls (P < 0.01). Spontaneous synaptic currents (SSCs) were also significantly increased in SCGNs within the co-cultured population of cells (P < 0.01). These SSCs could be blocked by a specific nicotinic receptor blocker, but not by a glutamate receptor antagonist. These observations clearly demonstrate the functional synapses formation increased in our co-culture system, suggesting that the “hybrid” synapse formation occurred between somatic and automatic neurons. Our data also indicate that acetylcholine and nicotinic receptors may be involved in mediating these processes.